Department of Orthopaedics, China-Japan Union Hospital of Jilin University, Changchun 130033, China.
Department of Joint Surgery of Orthopaedic Center, The Second Hospital of Jilin University, Changchun 130041, China.
Biomed Pharmacother. 2019 Oct;118:109395. doi: 10.1016/j.biopha.2019.109395. Epub 2019 Aug 29.
Osteosarcoma is one of malignant cancer. Histone phosphorylation is common in tumors. We explored the effects of p300-CBP-associated factor (PCAF) and phosphorylation of H3S28 in osteosarcoma cancer cell autophagy.
Osteosarcoma cancer cell lines were collected and/or transfected with full length PCAF or interference miRNAs to mimic or silence of PCAF expression. Immunoprecipitation assay and GST pull down was used to target targeting PCAF or H3S28ph. H3-/- SNU-C1 cells were transfected with H3WT- or H3S28F-expressing or enhanced green fluorescent protein (EGFP)-tagged LC3 plasmids, in which H3 was tagged with HA. An in vitro kinase activity assay was performed to test whether recombinant full-length PCAF could phosphorylate H3 in the site of S28. The functions on autophagy was detected by number of autophagosomes, number of EGFP-LC3, LC3-II/I, percentage of degradation and expression of autophagy associated gene (ATG).
PCAF positively regulated H3S28ph in osteosarcoma cancer cells; Immunoprecipitation assay and GST pull down demonstrated that PCAF could interact directly with H3 in osteosarcoma cancer cells. In addition, silence of PCAF inhibited the number of autophagosomes, number of EGFP-LC3, LC3-II/I, percentage of degradation and expression of ATG. Moreover, H3S28A (H3S28 mutation) impaired the promoting autophagy effects of PCAF. The PCAF-H3S28ph axis promoted osteosarcoma cancer autophagy viatranscriptional regulation of ATG genes.
PCAF regulated H3S28 phosphorylation and their axis promotes autophagy in osteosarcoma cancer cells viatargeting ATG5 and ATG7.
骨肉瘤是一种恶性肿瘤。组蛋白磷酸化在肿瘤中很常见。我们探讨了 p300-CBP 相关因子（PCAF）和 H3S28 磷酸化在骨肉瘤癌细胞自噬中的作用。
收集骨肉瘤癌细胞系，并/或用全长 PCAF 或干扰 miRNA 转染，以模拟或沉默 PCAF 的表达。免疫沉淀和 GST 下拉用于靶向 PCAF 或 H3S28ph。将 H3-/-SNU-C1 细胞转染表达 H3WT-或 H3S28F-或增强型绿色荧光蛋白（EGFP）标记的 LC3 质粒，其中 H3 被标记为 HA。进行体外激酶活性测定，以测试重组全长 PCAF 是否可以在 S28 位点磷酸化 H3。通过自噬体数量、EGFP-LC3 的数量、LC3-II/I 的百分比、降解百分比和自噬相关基因（ATG）的表达来检测自噬的功能。
PCAF 正向调节骨肉瘤癌细胞中的 H3S28ph；免疫沉淀和 GST 下拉表明 PCAF 可以在骨肉瘤癌细胞中与 H3 直接相互作用。此外，沉默 PCAF 抑制自噬体数量、EGFP-LC3 的数量、LC3-II/I 的百分比、降解百分比和 ATG 的表达。此外，H3S28A（H3S28 突变）削弱了 PCAF 促进自噬的作用。PCAF-H3S28ph 轴通过转录调控 ATG 基因促进骨肉瘤癌细胞自噬。
PCAF 调节 H3S28 的磷酸化，其轴通过靶向 ATG5 和 ATG7 促进骨肉瘤癌细胞自噬。
